As a highly-used model organism, Caenorhabditis elegans (C. elegans) worms offer a wide range of possibilities for scientific research. With advantages including, but not limited to, easy maintenance and handling, forward and reverse genetic manipulability, and a fully mapped nervous system connectome of 302 neurons, C. elegans offer a high throughput alternative to larger rodent model organisms. During the last decades C. elegans has become an indispensable tool for advanced research in the fields of aging, neurodegeneration, toxicology, drug discovery, and RNAi screening, to mention only a few. Attractive instruments for studying the function of C. elegans' nervous system are (i) worm trackers, i.e., software packages designed to quantify different aspects of C. elegans' behavior, (ii) monitoring neural activity using genetically encoded calcium indicators (calcium imaging), and (iii) manipulation of neural activity using optogenetics and femtosecond laser nanosurgery. However, because of technical shortcomings and lack of integration, in their current form these instruments cannot be considered adequate for next-generation research in various fields such as aging, neurodegeneration, toxicology, drug discovery, and RNAi screening applying C. elegans. In response to this challenge, this project aims to develop an innovative microscope system for sophisticated investigations of C. elegans' nervous system in action (Worm Investigator). The Worm Investigator system will allow scientists - for the first time - to monito and manipulate the nervous system of a freely moving worm using calcium imaging, optogenetics, and femtosecond laser nanosurgery while simultaneously tracking and analyzing the worm's complex behavior before, during, and after manipulation. This new commercial system will be based on four distinct innovations, to be developed as new embodiments of the following technology patented by MBF: (i) a dual-objective microscope setup; (ii) two Worm Tracking Objective Lens(tm) (WTOL) devices; (iii) a novel side ring LED illuminator (patent application in preparation); and (iv) our WormLab(r) worm tracking software. The benefit for the research community - and society in general - will be progress in next-generation research in the fields of aging, neurodegeneration, toxicology, drug discovery, and RNAi screening applying C. elegans. This will result in the ability to make new discoveries based on new studies that are currently not feasible which may ultimately provide the basis for novel treatment strategies for a wide variety of diseases. During Phase I we will develop a prototype dual- WTOL, further develop a novel side ring LED illuminator (part of our pre-existing technology), and develop a calibration test slide serving the specific needs of the Worm Investigator dual-WTOL microscope, as well as software to simultaneously move two WTOL devices and acquire synchronized images from each camera of the dual-WTOL microscope.